Calcium pyrophosphate dihydrate (CPPD) deposition disease is a common form of degenerative arthritis preferentially targeting the elderly. Affected patients have a severe chronic arthritis for which there are currently no therapies. As the population ages, CPPD deposition disease and other forms of degenerative arthritis will present an increasing societal burden in terms of health care and quality of life costs. CPPD crystals form in articular fibro-and hyaline cartilage. Their formation requires the interaction of extracellular pyrophosphate and calcium in a milieu of disordered pericellular matrix. The contributions of cartilage extracellular matrix to CPPD crystal formation remain poorly characterized and are the focus of this work. We identified the transglutaminase (Tgase) family of enzymes as important participants in CPPD crystal formation. Tgases catalyze the formation of unique crosslinks between or within proteins. Articular cartilage contains two Tgase enzymes, type II Tgase and Factor XIIIA (FXIIIA). Tgase activity dramatically increases with age in articular cartilage, and is present in the pericellular chondrocyte matrix at sites of potential crystal formation. Inhibition of Tgase activity suppresses CPPD crystal formation in vitro, while Tgase overexpression causes matrix mineralization. We hypothesize that increased Tgase activity in the extracellular matrix of aging cartilage contributes to CPPD crystal formation by crosslinking key CPPD crystal-promoting proteins into pericellular matrix. We will show that 1) extracellular matrix Tgase activity participates in CPPD crystal formation; 2) FXIIIA is the predominant Tgase in cartilage extracellular matrix; and 3) three key Tgase substrates (osteopontin, SPARC, and fibrillin 1) are trapped in the pericellular matrix by Tgase and directly participate in CPPD crystal formation. We will explore these hypotheses by using porcine cartilage from animals of three age groups, and human cartilage from patients with CPPD deposition disease or osteoarthritis. We will use two models of CPPD crystal formation to investigate the role of Tgase and its substrate proteins in CPPD crystal formation. Immunohistochemical studies of diseased human cartilage will be used to confirm the relevance of these findings to CPPD disease. These studies will contribute to our understanding of CPPD deposition disease, so that effective therapies for this common and disabling form of arthritis can be designed.